Circadian and photoperiodic regulation of the vegetative to reproductive transition in plants

As sessile organisms, plants must respond constantly to ever-changing environments to complete their life cycle; this includes the transition from vegetative growth to reproductive development. This process is mediated by photoperiodic response to sensing the length of night or day through circadian regulation of light-signaling molecules, such as phytochromes, to measure the length of night to initiate flowering. Flowering time is the most important trait to optimize crop performance in adaptive regions. In this review, we focus on interplays between circadian and light signaling pathways that allow plants to optimize timing for flowering and seed production in Arabidopsis, rice, soybean, and cotton. Many crops are polyploids and domesticated under natural selection and breeding. In response to adaptation and polyploidization, circadian and flowering pathway genes are epigenetically reprogrammed. Understanding the genetic and epigenetic bases for photoperiodic flowering will help improve crop yield and resilience in response to climate change.

The manuscript contains a large number of references and I have no major concerns with the validity of the statements made about gene function and the interactions between various components of these pathways.However, I found the structure of the paper and the areas of focus to be unusual and made the paper, for me, difficult to follow.In sections, the manuscript read as a list of statements about gene functions and lacked a strong narrative that tied all this information together.This may have been the intention of the authors and for their target audience, just that I found the manuscript quite challenging to follow.Photoperiodism is such a large area of research that I think a review would benefit from greater focus and a more logical structure.In addition to the lack of a single narrative flow, basic concepts of photoperiodism (e.g.short day and long day requirements to flower, circadian gating) are left undescribed even at a high level, in place of extremely detailed descriptions of the interactions between a great number of genes.Information is grouped unusually, with unrelated topics included in the same sections and other sections that do not have an obvious link to photoperiodism (such as cotton evolution) which might be better described in a different manuscript.Some specific comments on the structure: -Photoperiodism and circadian regulation affects many developmental processes, but it should be clear in the introduction, abstract and title that the focus of the review is on the vegetative to reproductive transition.The current title is very broad and the introduction section mentions different biological processes that are not relevant to the current study.A greater focus on the vegetative to reproductive transition would clarify for the reader what the focus of the paper is.
-The introduction section is potentially unnecessary and describes a lot of information on genes and their potential roles that are later repeated in specific sections.-Many concepts are not described adequately.It is not stated that Arabidopsis is a long-day plant and that these pathways have adapted to different photoperiods to promote flowering.The concept of circadian gating is not described, nor the temperature-mediated changes in the clock.-There is a section combining epistasis and epigenetics, two different topics, that are covered very briefly with few examples and do not seem to be especially important for photoperiodism (These are facets across gene regulation).The inclusion of epistasis in this section is particularly unusual.The section then moves to description of polyploidization in cotton and a description of genes in this species.I understand this is an area of research expertise for the authors, but the topic here does not have special interest for photoperiodism and should probably be left out.
-The concluding remarks section mentions that an understanding of these pathways can lead to engineering yield improvement.It would be good to include some examples of crops for which this has been achieved to demonstrate the real-world outcomes of this research area.Overall, I feel that the manuscript could be improved by clearly defining the topic and target audience, then rewriting to focus on these topics so they are most clearly understood, with a grounding in some of the underlying concepts.This will help readers to understand the relevance of each gene pathway and the broader context of how they fit in to plant biology.

Minor comments
Line 41: Replace "Despite" with "although.Line 42: "key regulators of floral transition are highly conserved".This is poorly defined.I am familiar with work from wheat that shows the CONSTANS proteins, for example, have a less important role in defining flowering time in this species.I believe this sentence should either be modified or removed.Line 84 -Sentence is incomplete, ends on "constitute".Lines 113-116: These sentences seem out of place since they describe the conservation of regulators in different species.Was this the topic of the introduction?Line 203: "switch on flowering" could be defined in more technical terms.Lines 238 -242: The genes mentioned here are from Soybean.Why is this information not grouped together in the section describing responses in soybean?Line 246: The evening complex in Arabidopsis is introduced mid-paragraph.

Point-by-point response
Reviewer #1 (Remarks to the Author): The review by Wang et al. provides a comprehensive description of the genetic pathways underlying photoperiodism and circadian responses the vegetative to reproductive transition in different plant species.This is an important topic that influences crop adaptation to different environments and has been the subject of much genetic research which is described in this review.The manuscript contains a large number of references and I have no major concerns with the validity of the statements made about gene function and the interactions between various components of these pathways.However, I found the structure of the paper and the areas of focus to be unusual and made the paper, for me, difficult to follow.In sections, the manuscript read as a list of statements about gene functions and lacked a strong narrative that tied all this information together.This may have been the intention of the authors and for their target audience, just that I found the manuscript quite challenging to follow.Photoperiodism is such a large area of research that I think a review would benefit from greater focus and a more logical structure.In addition to the lack of a single narrative flow, basic concepts of photoperiodism (e.g.short day and long day requirements to flower, circadian gating) are left undescribed even at a high level, in place of extremely detailed descriptions of the interactions between a great number of genes.Information is grouped unusually, with unrelated topics included in the same sections and other sections that do not have an obvious link to photoperiodism (such as cotton evolution) which might be better described in a different manuscript.
Response: We thank this expert reviewer for the insightful analysis and constructive comments to improve the paper.Our responses to specific comments are as follows.
Some specific comments on the structure: -Photoperiodism and circadian regulation affects many developmental processes, but it should be clear in the introduction, abstract and title that the focus of the review is on the vegetative to reproductive transition.The current title is very broad and the introduction section mentions different biological processes that are not relevant to the current study.A greater focus on the vegetative to reproductive transition would clarify for the reader what the focus of the paper is.In the revision, we added paragraphs in the Introduction to clarify basic concepts such as short-day and long-day requirements for flowering, as well as circadian gating, to provide a clear context for readers.Additionally, we have reorganized the relevant information to focus more closely on the genetic pathways underlying photoperiodism and circadian responses.Some unrelated topics and subjects including list of genes and their potential roles have been removed to enhance the clarity and coherence of the paper.
-Many concepts are not described adequately.It is not stated that Arabidopsis is a long-day plant and that these pathways have adapted to different photoperiods to promote flowering.The concept of circadian gating is not described, nor the temperature-mediated changes in the clock.

Response:
We have revised the manuscript to address these issues.We have introduced the concepts of long-day, short-day, and day-neutral plants, clarifying that Arabidopsis is a facultative long-day plant with metabolic pathways adapted to different photoperiods for flowering promotion.Additionally, we have provided conceptual explanation of circadian gating and discussed temperature-mediated changes in the clock to ensure a more comprehensive coverage of the topic.
-There is a section combining epistasis and epigenetics, two different topics, that are covered very briefly with few examples and do not seem to be especially important for photoperiodism (These are facets across gene regulation).The inclusion of epistasis in this section is particularly unusual.The section then moves to description of polyploidization in cotton and a description of genes in this species.I understand this is an area of research expertise for the authors, but the topic here does not have special interest for photoperiodism and should probably be left out.
Response: We agree in principle with this assessment.Our intention was to make the readers aware of complicated gene and allelic interactions in complex crop genomes, in addition to transcriptional regulation by DNA methylation and chromatin modifications in response to seasonal changes and environmental cues.Compared to detailed studies in Arabidopsis, the underlying mechanisms and pathways are much less known in crop plants, many of which are polyploids or of polyploid origin.To facilitate a smooth transition, we added a paragraph to expand our understanding of epialleles such as FLC and FWA; the former is involved in response to cold winder via a vernalization process.Many other factors such CO in cotton are epialleles, which result from intergenomic interactions in polyploid crops.The impact of epistasis is much less understood due to duplicate genes and networks in polyploid plants and crops.We hope the coverage of these topics will help introduce and develop translational research on crop plants.Although the findings are relatively limited compared to deep understanding in Arabidopsis, they should provide some guidelines for future studies on translating basic findings in Arabidopsis to crop improvement.
-The concluding remarks section mentions that an understanding of these pathways can lead to engineering yield improvement.It would be good to include some examples of crops for which this has been achieved to demonstrate the real-world outcomes of this research area.
Response: Thank you for the suggestion."There are good examples of the genes in the circadian clock and photoperiodic flowering pathways that are associated with agronomic traits in crops.Rice Hd1 1 , an ortholog of the Arabidopsis CO, controls expression of two florigen genes Hd3a and RFT1 2-4 during the floral transition.ZmCCT9 in maize is diurnally regulated and negatively regulates the expression of the florigen ZCN8 through a CACTAlike TE insertion in ZmCCT promoter region 5 , resulting in reduction of photoperiod sensitivity, thus accelerating maize spread to long-day environments 6 .The circadian clock of cultivated tomatoes has slowed during domestication.EMPFINDLICHER IM DUNKELROTEN LICHT1 (EID1) is an F-box protein that targets phytochrome A for degradation in Arabidopsis 7 .The EID1 allele in cultivated tomatoes is under selection sweep and enhances plant performance under long day photoperiods presumably resulting from moving away from the equator 8 .Fruit yield in hybrid tomato is related to SINGLE FLOWER TRUSS (SFT) 9 , a homolog of FT in Arabidopsis.Moreover, epigenetic alteration of circadian clock genes including CCA1 and LHY in plant hybrids are related to growth vigor in Arabidopsis 10-12 and maize 13,14 .In cotton, GhCOL2 is an epiallele, which may be selected during domestication to increase its expression and thereby reduce photoperiod sensitivity, helping spread worldwide cotton cultivation 15 .These examples will help us design strategies to optimize circadian input and output signals and improve flowering time and crop yield using genome-and gene-editing tools."This is included in Conclusion.
Overall, I feel that the manuscript could be improved by clearly defining the topic and target audience, then rewriting to focus on these topics so they are most clearly understood, with a grounding in some of the underlying concepts.This will help readers to understand the relevance of each gene pathway and the broader context of how they fit in to plant biology.
Response: Thank you for your constructive comments.We have revised the manuscript to concentrate on the regulation of plant transitions from vegetative to reproductive stages, specifically highlighting photoperiodism, circadian rhythm, and epigenetic regulation.We have included additional context in the Introduction, Crop Flowering Time, and Conclusions to facilitate a clearer understanding and transition of these topics.

Minor comments
Line 41: Replace "Despite" with "although.Response: The sentence along with other context was removed in the revised Introduction.
Line 42: "key regulators of floral transition are highly conserved".This is poorly defined.I am familiar with work from wheat that shows the CONSTANS proteins, for example, have a less important role in defining flowering time in this species.I believe this sentence should either be modified or removed.Response: As suggested, the sentence was removed.
Line 84 -Sentence is incomplete, ends on "constitute".Response: We have completed the sentence as "…whose expression peaks in the evening; they constitute the central loop of the core oscillator." Lines 113-116: These sentences seem out of place since they describe the conservation of regulators in different species.Was this the topic of the introduction?Response: We moved these sentences to the top of the paragraph under the section of Photoperiodic Control of Flowering in Crop/Rice.Line 203: "switch on flowering" could be defined in more technical terms.Response: We replaced "switch on flowering" with "trigger flowering".
Lines 238 -242: The genes mentioned here are from Soybean.Why is this information not grouped together in the section describing responses in soybean?Response: Sorry for the confusion.We moved the first sentence to the section of soybean, while the sorghum PRR37 remains in the same place, followed by, "Similarly, clock components such as OsPRR37/DTH7 and the EC components, also regulate rice flowering…" Line 246: The evening complex in Arabidopsis is introduced mid-paragraph.Response: We started the EC complex in a new paragraph.

Figure 2 :
What are the line graphs at the bottom of the figure?Reviewer #2 (Remarks to the Author): This submission by Wang et al. provides a timely and comprehensive overview of the molecular mechanisms underlying photoperiodic regulation of flowering times in the model plant Arabidopsis thaliana and several economically-important crops.Overall the manuscript is well written.Below are suggestions for improvement.1. Title: it would be better to remove the word of crops because crops are plants.2. Lines 43-58: As these two paragraphs describe the mechanisms in Arabidopsis thaliana, not in other plants, please add the species name somewhere in the first paragraph.3. Line 84: ……constitute?4. Lines 312-317: the function of COOLAIR in vernalization-mediated FLC repression has been questioned (e.g.DOI: 10.7554/eLife.84594and other publications).Please rephrase this paragraph.5.A few typos.Line 91: actives to activates.Line 114: …conserved IN duckweed… Line 135: diel to diurnal?Line 311: …cold IN winter… Line 203: please italicize OsMADS14 and OsMADS15.6. Lines 318-320: these two sentences are not consistent.

Response:
As suggested, we have revised the title as well as the structure of the manuscript to provide a clearer narrative flow and better organization of topics related to photoperiodism and circadian responses.The new title is "Photoperiodism and Circadian Regulation from Vegetative to Reproductive Transition in Plants" to better highlight the focus on the transition from vegetative growth to reproduction.

Figure 2 :
Figure 2: What are the line graphs at the bottom of the figure?Response: Sorry for the confusion.The line graphs at the bottom of the figure 2 indicate mRNA oscillation of CO, CO/FKBP12, and FT, respectively.We clarified this in the legend and also in the text.